gaughan at bms.com
Tue Nov 21 11:22:15 EST 1995
In article <48sqqd$jvs at synapse.bms.com>, Scott Taylor <taylor_st at bms.com> wrote:
> Does anyone know of vendors which sell a machine which
> allow you to add one ml of cell culture in and get qood quality
> DNA out (good enough for automated sequencing)?
Depends on your budget. We had a look at the Vistra Labstation. It will take
cultures thru plasmid preps and then thru sequencing reactions. (At the time
we looked at it one still had to do the last step of dye terminator removal
but they thought they'd be able to add that step as well in the near
future.) It worked reasonably well. Negatives: It took ca. 5 - 6 hr to
prepare 16 samples
(should have been upgraded by now to do 24) so one could not get a full
gel's worth of samples ready for same day loading. Quality of sequence was
OK but not as good as 'manual' - possible sources of loss of quality are
template and less good thermal control in microtiter plate thermocycler.
Loss of quality not dramatic - a few % more N's over the 1st 350 bases or
so (we only looked at plasmid templates - assume M13 gives longer reads.
We also looked at this before ThermoSequenase and Tag fs became
available). Cost was 50-60k$. An added benefit was that the machine could
also be programmed to do e.g. plasmid preps & restriction digests and
other useful chores. At the low budget end we looked at McConnell
Research's gel-based instrument (template prep only). They recommend an
extra purification step (pptn) for automated sequencing templates; we got
good results diluting and reconcentrating w a microcon 100 (more expensive
but less time demanding and less prone to operator error). Yeilds of
plasmid were significantly less than with 'manual' methods. (McConnell
provides a special growth medium & fairly explicit protocols for culture -
we used these but still got less than good yeilds.) This was a problem if
you have many reaxns to do on a single template. Quality of results (after
microcon) were very good.
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